The goal of this application is to investigate the feasibility and requirements of converting existing in vivo, differentiated cells of the adult CNS - such as astroglia - into functional and appropriately integrated neurons, using in situ directed neuronal conversion (IDNC). IDNC is novel and distinct from stem cell approaches. With IDNC, a cocktail of transcription regulators is transduced into mature CNS cells, such as astroglia, using polycistronic lentiviral vectors or otherwise. These factors lead to a rapid and directed conversion into a distinct mature neuronal phenotype. Such directed conversion challenges the basic tenet that differentiated cell phenotypes are inherently stable - particularly in situ in the CNS. Directed conversion potentially overcomes some of the inherent difficulties with other therapeutic cell replacement strategies, such as stem cell approaches. Directly converting existing cells may be least intrusive on the general architecture of existing circuitry. A second and perhaps greater challenge is the functional integration of newly converted neurons into CNS local circuitry. This problem will be address using a variety of physiological and optical imaging approaches. PUBLIC HEALTH RELEVANCE: The generation of new CNS neurons that appropriately integrate into the mature mammalian CNS represents a major challenge in regenerative medicine. We present a novel and conceptually simple approach: to generate new mammalian CNS neurons in situ by in vivo directed conversion of other CNS cells. We will determine whether such cells can integrate into the existing neuronal circuitry.